bims-istrec 2021-10-03 papers

bims-istrec

Biomed News

on Integrated stress response in cancer

Issue of 2021–10–03
eight papers selected by
the Vincenzo Ciminale lab, Istituto Oncologico Veneto

Human β-defensin 1 Functions as a Tumor Suppressor via ER Stress-triggered JNK pathway in Hepatocellular Carcinoma.
MFN2 Stabilization: A Bridge for Endoplasmic Reticulum Stress Sensitivity in Melanoma.
Salubrinal Enhances Cancer Cell Death during Glucose Deprivation through the Upregulation of xCT and Mitochondrial Oxidative Stress.
Fisetin Protects HaCaT Human Keratinocytes from Fine Particulate Matter (PM2.5)-Induced Oxidative Stress and Apoptosis by Inhibiting the Endoplasmic Reticulum Stress Response.
Cytotoxicity of 4-hydroxy-N-(naphthalen-1-yl)-2-oxo-2H-chromene-3-carboxamide in multidrug-resistant cancer cells through activation of PERK/eIF2α/ATF4 pathway.
6-Methoxylated Flavonoids: Jacein, and 3-demethyljacein from Centaurea schmidii with Their Endoplasmic Reticulum Stress and Apoptotic Cell Death in Breast Cancer Cells Along with In-silico Analysis.
Marchantia polymorpha L. ethanol extract induces apoptosis in hepatocellular carcinoma cells via intrinsic- and endoplasmic reticulum stress-associated pathways.
Salubrinal Regulates the Apoptosis of Adrenocortical Carcinoma Cells via the PERK/eIF2α/ATF4 Signaling Pathway.

J BUON. 2021 Jul-Aug;26(4):26(4): 1365-1372

Human β-defensin 1 Functions as a Tumor Suppressor via ER Stress-triggered JNK pathway in Hepatocellular Carcinoma.

Xinhong Li, Wanying Song, Mingyu Zhang, Pengwei Zhao.

PURPOSE: Human β-defensin 1 (DEFB1) belongs to defensins family that contribute to innate immune responses and was recently found to downregulate a variety of cancers, including renal, prostatic, and oral squamous cell carcinoma, and therefore is considered as a potential tumor suppressor. However, the role of DEFB1 in hepatocellular carcinoma (HCC) still needs to be elucidated.
METHODS: Quantitative PCR and Western blot were used to measure the expression levels of interested proteins. CCK-8 and colony formation assays were performed to determine the ability of cell proliferation. Tumor formation experiments in nude mice were used to examine the tumor growth.
RESULTS: The expression level of DEFB1 was dramatically downregulated in human HCC. Quantitative PCR and Western blot results also showed a pronounced decrease of DEFB1 expression in the liver cancer cell lines. Rescuing the expression of DEFB1 in Huh7 cells effectively suppressed cell proliferation and reduced the colony forming ability, probably by inducing cell apoptosis and cell cycle arrest. Moreover, tumor formation experiments in nude mice also showed inhibition of tumor growth by DEFB1 expression in vivo. Furthermore, induction of DEFB1 expression induced degraded protein increase and endoplasmic reticulum (ER) stress, which subsequently activated JNK pathway. Pharmacologic inhibition of ER stress by 4-phenylbutyrate, a compound to alleviate ER stress, effectively eliminated DEFB1-induction inhibition of cell proliferation and migration.
CONCLUSION: DEFB1 functions as a tumor suppressor in HCC through activating ER stress and JNK pathway, which may provide a potential strategy for HCC treatment.
J Invest Dermatol. 2021 Sep 23. pii: S0022-202X(21)01224-0. [Epub ahead of print]

MFN2 Stabilization: A Bridge for Endoplasmic Reticulum Stress Sensitivity in Melanoma.

Charles H Adelmann, Inbal Rachmin, David E Fisher.

In a new article in the Journal of Investigative Dermatology, Wang et al. (2021) report that mitochondrial quality control modulates responses to endoplasmic reticulum (ER) stress in melanoma. They implicate a linear pathway of XBP1, MARCH5, and MFN2 that act together to regulate mitochondrial fission and mitophagy and ultimately mediate melanoma cell sensitivity to ER stress. This work informs therapeutic combinations and biomarker strategies for targeting melanoma organellar homeostasis as well as for life‒death decisions.

DOI: https://doi.org/10.1016/j.jid.2021.05.003
Biomedicines. 2021 Aug 28. pii: 1101. [Epub ahead of print]9(9):

Salubrinal Enhances Cancer Cell Death during Glucose Deprivation through the Upregulation of xCT and Mitochondrial Oxidative Stress.

Mei-Chun Chen, Li-Lin Hsu, Sheng-Fan Wang, Yi-Ling Pan, Jeng-Fan Lo, Tien-Shun Yeh, Ling-Ming Tseng, Hsin-Chen Lee.

  Cancer cells have the metabolic flexibility to adapt to heterogeneous tumor microenvironments. The integrated stress response (ISR) regulates the cellular adaptation response during nutrient stress. However, the issue of how the ISR regulates metabolic flexibility is still poorly understood. In this study, we activated the ISR using salubrinal in cancer cells and found that salubrinal repressed cell growth, colony formation, and migration but did not induce cell death in a glucose-containing condition. Under a glucose-deprivation condition, salubrinal induced cell death and increased the levels of mitochondrial reactive oxygen species (ROS). We found that these effects of salubrinal and glucose deprivation were associated with the upregulation of xCT (SLC7A11), which functions as an antiporter of cystine and glutamate and maintains the level of glutathione to maintain redox homeostasis. The upregulation of xCT did not protect cells from oxidative stress-mediated cell death but promoted it during glucose deprivation. In addition, the supplementation of ROS scavenger N-acetylcysteine and the maintenance of intracellular levels of amino acids via sulfasalazine (xCT inhibitor) or dimethyl-α-ketoglutarate decreased the levels of mitochondrial ROS and protected cells from death. Our results suggested that salubrinal enhances cancer cell death during glucose deprivation through the upregulation of xCT and mitochondrial oxidative stress.

Keywords:  integrated stress response; oxidative stress; salubrinal; xCT

DOI:  https://doi.org/10.3390/biomedicines9091101
Antioxidants (Basel). 2021 Sep 18. pii: 1492. [Epub ahead of print]10(9):

Fisetin Protects HaCaT Human Keratinocytes from Fine Particulate Matter (PM2.5)-Induced Oxidative Stress and Apoptosis by Inhibiting the Endoplasmic Reticulum Stress Response.

Ilandarage Menu Neelaka Molagoda, Mirissa Hewage Dumindu Kavinda, Yung Hyun Choi, Hyesook Lee, Chang-Hee Kang, Mi-Hwa Lee, Chang-Min Lee, Gi-Young Kim.

  Fine particulate matter (PM2.5) originates from the combustion of coal and is found in the exhaust of fumes of diesel vehicles. PM2.5 readily penetrates the skin via the aryl hydrocarbon receptor, causing skin senescence, inflammatory skin diseases, DNA damage, and carcinogenesis. In this study, we investigated whether fisetin, a bioactive flavonoid, prevents PM2.5-induced apoptosis in HaCaT human keratinocytes. The results demonstrated that fisetin significantly downregulated PM2.5-induced apoptosis at concentrations below 10 μM. Fisetin strongly inhibited the production of reactive oxygen species (ROS) and the expression of pro-apoptotic proteins. The PM2.5-induced apoptosis was associated with the induction of the endoplasmic reticulum (ER) stress response, mediated via the protein kinase R-like ER kinase (PERK)-eukaryotic initiation factor 2α (eIF2α)-activating transcription factor 4 (ATF4)-CCAAT-enhancer-binding protein (C/EBP) homologous protein (CHOP) axis. Additionally, the cytosolic Ca2+ levels were markedly increased following exposure to PM2.5. However, fisetin inhibited the expression of ER stress-related proteins, including 78 kDa glucose-regulated protein (GRP78), phospho-eIF2α, ATF4, and CHOP, and reduced the cytosolic Ca2+ levels. These data suggest that fisetin inhibits PM2.5-induced apoptosis by inhibiting the ER stress response and production of ROS.

Keywords:  PM2.5; apoptosis; endoplasmic reticulum stress; fisetin; reactive oxygen species

DOI:  https://doi.org/10.3390/antiox10091492
Biochem Pharmacol. 2021 Sep 25. pii: S0006-2952(21)00404-4. [Epub ahead of print] 114788

Cytotoxicity of 4-hydroxy-N-(naphthalen-1-yl)-2-oxo-2H-chromene-3-carboxamide in multidrug-resistant cancer cells through activation of PERK/eIF2α/ATF4 pathway.

Xiaohua Lu, Ge Yan, Sabine M Klauck, Edmond Fleischer, Anette Klinger, Yoshikazu Sugimoto, Letian Shan, Thomas Efferth.

  After decades of research, multidrug resistance (MDR) remains a huge challenge in cancer treatment. In this study, the cytotoxic of 4-hydroxy-N-(naphthalen-1-yl)-2-oxo-2H-chromene-3-carboxamide (MCC1734) has been investigated towards multidrug-resistant cancer cell lines. MCC1734 exerted cytotoxicity on cell lines expressing different mechanisms of drug resistance (P-glycoprotein, BCRP, ABCB5, EGFR, p53 knockout) to a different extent. Interestingly, sensitive CCRF-CEM cells and multidrug-resistant P-gp-overexpressing CEM/ADR5000 cells represented similar sensitivity towards MCC1734, indicating MCC1734 can bypass P-gp-mediated resistance. Microarray-based mRNA expression revealed that MCC1734 affected cells by multiple pathways, including cell cycle regulation, mitochondrial dysfunction, apoptosis signaling, and EIF2 signaling. MCC1734 stimulated the generation of excessive reactive oxygen species and the collapse of mitochondria membrane potential in CCRF-CEM cells, companied by the arrest of the cell cycle in the G2M phase and apoptosis induction as determined by flow cytometry. In addition, our immunoblotting analysis highlighted that MCC1734 triggered endoplasmic reticulum (ER) stress, evidenced by the activation of p-PERK, p-eIF2α, ATF4 and CHOP. The anti-cancer effects of MCC1734 were further observed in vivo using human xenograft tumors transplanted to zebrafish, providing further support for MCC1734 as a promising new candidate for cancer drug development.

Keywords:  Apoptosis; Cancer; Drug development; ER stress; Multidrug resistance; PERK/eIF2α/ATF4 pathway

DOI:  https://doi.org/10.1016/j.bcp.2021.114788
Iran J Pharm Res. 2021 ;20(2): 417-432

6-Methoxylated Flavonoids: Jacein, and 3-demethyljacein from Centaurea schmidii with Their Endoplasmic Reticulum Stress and Apoptotic Cell Death in Breast Cancer Cells Along with In-silico Analysis.

Mahmoud Aghaei, Mahmoud Mirzaei, Mustafa Ghanadian, Moslem Fallah, Roodabeh Mahboodi.

  In phytochemical analysis, Jacein derivatives: 5,7,4'-trihydroxy-3,6,3'-trimethoxyflavone-7(β)-D-glucopyranoside (1), and 3-demethyljacein: 3,5,7,4'-tetrahydroxy-6,3'-dimethoxyflavone-7(β)-D-glucopyranoside (2) were isolated from Campylopus schmidii (C. schmidii) for the first time. The structures were determined by interpretation of NMR, UV, and Mass spectra. To check the roles of ER stress and consequent apoptosis in MCF-7 cell by these compounds, UPR signaling pathway was further examined by analysis of expression of ER stress-related genes. In MTT assay, compounds 1-2 showed cytotoxicity activity against MCF-7 (A) and MDA-MB cells (B) with IC50 values (μM) of 1) 60.04 ± 7.98 (A), and > 200 (B); 2) 42.89 ± 1.91 (A), and 85.31 ± 2.68 (B). The Annexin/PI flow cytometry apoptosis of tested compounds 1-2 was increased significantly in a dose-dependent manner. For example, MCF-7 treatment at the concentration of 100 μM of compounds 1, 2 resulted in total apoptosis (early + late) of 42.04 (18.1 + 24.0), and 66.49 (2.7 + 63.8)%, respectively. Fluorescence microscopy analysis detected an increased protein aggregation, indicating induced ER stress with a marked increase in XBP-1, sXBP-1, ATF-4, and CHoP compared to untreated cells. In-silico characterization, suggested that Adenosine diphosphate site (A-site) and quercetin site (Q-Site) in IRE1a enzyme are both available interacting sites of a target for the investigated ligands but with different strengths of interactions. The results indicated that the ligand∼A-Site complexes are stronger than the ligand∼Q-Site complexes, but the already available ADP ligand in cells does not allow other ligands to interact with the A-Site and cause them to bond in Q-Site.

Keywords:  Apoptosis; Breast cancer; Centaurea; Endoplasmic reticulum stress; Flavonoid; In-silico analysis

DOI:  https://doi.org/10.22037/ijpr.2020.113895.14548
Chin Med. 2021 Sep 28. 16(1): 94

Marchantia polymorpha L. ethanol extract induces apoptosis in hepatocellular carcinoma cells via intrinsic- and endoplasmic reticulum stress-associated pathways.

Fangfang Zhou, Adila Aipire, Lijie Xia, Xierenguli Halike, Pengfei Yuan, Mamtimin Sulayman, Weilan Wang, Jinyao Li.

   BACKGROUND: Marchantia polymorpha L. is a kind of Chinese herbal medicine and has various biological activities including antioxidant and antifungal. However, it is not clear about the antitumor effect and mechanism of M. polymorpha. We prepared M. polymorpha ethanol extract (MPEE) and investigated its antitumor effect on hepatocellular carcinoma cells both in vitro and in vivo.
METHODS: The viability of hepatocellular carcinoma cells was detected by MTT assay. The distribution of cell cycle was analyzed by propidium iodide (PI) staining. The morphology of nuclei was observed by Hoechst 33258 staining. Apoptosis was detected by Annexin V/PI staining. JC-1 fluorescent probe and DCFH-DA were used to detect the mitochondrial membrane potential (ΔψM) and the level of reactive oxygen species (ROS), respectively. Caspase inhibitors were used to test the function of caspase in the induction of apoptosis. Quantitative real time polymerase chain reaction (qRT-PCR) and Western blot were used to evaluate the levels of mRNA and protein, respectively. Differentially expressed genes and signaling pathways were identified by transcriptome analysis. The H22 tumor mouse model was used to detect the antitumor effect of the extract.
RESULTS: MPEE significantly suppressed the migration and growth of BEL-7404, HepG2 and H22 cells in a dose- and time-dependent manner through induction of apoptosis characterized by chromosomal condensation and cell cycle arrest at G0/G1 and G2/M phases. MPEE induced mitochondria-dependent apoptosis via upregulation of Bax and downregulation of Bcl-2 to reduce mitochondrial membrane potential and increase the release of cytochrome c. The levels of cleaved caspase-8 and -9 were significantly increased, which sequentially activated caspase-3 to cleave PARP. We further found that MPEE significantly increased ROS production and activated endoplasmic reticulum (ER) stress associated-apoptotic signaling pathway. Moreover, MPEE significantly inhibited H22 tumor growth in mouse model and improved the survival of tumor mice.
CONCLUSION: These results suggested that MPEE suppressed hepatocellular carcinoma cell growth through induction of apoptosis via intrinsic- and ER stress-associated pathways.

Keywords:  Apoptosis; Hepatocellular carcinoma; Marchantia polymorpha L.; Signaling pathway; Tumor mouse model

DOI:  https://doi.org/10.1186/s13020-021-00504-4
Int J Endocrinol. 2021 ;2021 5038130

Salubrinal Regulates the Apoptosis of Adrenocortical Carcinoma Cells via the PERK/eIF2α/ATF4 Signaling Pathway.

Lili Wu, Chunfeng Liang, Xuemei Huang, Xiujun Deng, Jiming Jiang, Zuojie Luo.

The protein-kinase-R- (PKR-) like endoplasmic reticulum kinase (PERK) signaling pathway is a well-known promoter of cell apoptosis. In this study, we aimed to determine whether salubrinal (Sal), a selective activator of eukaryotic translation initiation factor 2 (eIF2α), can induce apoptosis of human adrenocortical carcinoma (ACC) cell via activating the PERK/eIF2α/ATF4 signaling pathway, and the potential mechanisms of this action were explored. The ACC cell lines, including SW-13 and NCI-H295 R, were used. 3-(4,5)-Dimethylthiazol(-z-y1)-3,5-di-phenytetrazoliumromide (MTT) assay, cell scratch experiments, flow cytometry, and JC-1 staining assays were performed to detect the cell viability, cell migration, and cell apoptosis. The expression of PERK/eIF2α/ATF4 signaling-pathway-related proteins and apoptosis-related proteins was detected by western blot (WB). Intracellular Ca2+ ion concentration was determined by a confocal laser scanning microscope. The results showed that Sal inhibited the migration and proliferation of ACC cells. Sal remarkably increased the influx of Ca2+ ion and the apoptosis rate of ACC cells in vitro. Furthermore, the expression levels of PERK/eIF2α/ATF4 signaling-related proteins and apoptosis-related proteins were upregulated in the treatment of Sal. The research demonstrated that Sal reduces the cell viability, increases the intracellular calcium concentration, and promotes the apoptosis of ACC cells in vitro through increasing the phosphorylation level of eIF2α and activating the PERK/eIF2α/ATF4 signaling. PERK/eIF2α/ATF4 is expected to act as a potential therapeutic target for the treatment of adrenocortical carcinoma.

DOI: https://doi.org/10.1155/2021/5038130